Matrix-Assisted Laser Desorption Ionization (MALDI) mass spectrometry is a bioanalytical mass spectrometry technique used to qualitatively analyze peptides and proteins. Fenselau, C., MALDI MS and Strategies for protein Analysis, Analytical Chemistry, 1997, 661a-5a. The technique has also been applied to technical polymers. Wu, K. J., Odom, R. W., Characterizing Synthetic, Analytical Chemistry, Jul. 1, 1998, 456. In MALDI, the sample and a small UV absorbing, Brönsted acidic molecule, the matrix, are dissolved in a solvent and co-crystallized. The matrix and sample are then irradiated with an ultraviolet laser. The matrix, which is in excess, with a molar ratio of 100-10,000:1 to the sample, absorbs the energy from the laser and protonates the sample molecule. Traditionally in mass spectrometry, the energy required to produce ionization of the sample molecule also causes fragmentation. But in MALDI, the matrix absorbs most of the energy from the laser and the sample molecule is desorbed intact. Therefore, the acquired mass spectrum contains few peaks; the molecular ion singly, doubly and triply charged (i.e. MH+, MH+2 and MH+3) as well as dimers and trimers that are also singly, doubly and triply charged.
The major disadvantage to MALDI is the difficulty in providing quantitative information due to matrix and sample sensitivity. Vesting, M. M., In Time-of-Flight Mass Spectrometry; Cotter, R. J.; American Chemical Society Symposium Series 549: Washington, D.C., 1994; 211-24.
In the experiments reported here calibration curves were constructed for human serum albumin, lactoferrin and lysozyme utilizing cytochrome C as an internal standard. The calibration curves were used to quantify the protein uptake in a biomaterial. The results were compared to an ultra-violet (UV) colorimetric assay. The UV and MALDI results were compared to each other and to theoretical values. The MALDI and UV results were within 15% of each other and theoretical values.
The method of this invention provides for elimination of calibration curves and the implementation of a simple equation. The validity of the equation was confirmed with further experiments utilizing carbonic anhydrase, myoglobin, αlactalbumin and horse albumin with cytochrome C as an internal standard.